With this proposal, we intend to test the overall hypothesis that type 4 cyclic nucleotide phosphodiesterases (PDE4s) are essential for cAMP homeostasis and receptor signaling specificity. There is ample pharmacological evidence that PDE4s contribute to the shaping of the cAMP signals arising form adrenergic, prostaglandin, and gonadotropin receptors. However, the function of individual PDE4 isoforms in a cell is largely unknown. The presence of four PDE4 genes and more than twenty splicing variants prompts the question of the functional significance of such a complexity. We propose that different PDE4 variants function in macromolecular complexes to specify signals that control most biological processes, including excitation/contraction coupling in cardiomyocytes. These complexes contribute to the divergent biological responses associated with 1 and 2 adrenergic receptors. Indeed, our analysis of PDE4 KO phenotypes has provided initial evidence that PDE4D and PDE4B play distinct functions in the body. A late onset cardiomyopathy associated with arrhythmias and cardiac failure after PDE4D ablation implies that removal of a single PDE4 produces subtle but critical losses of function in the heart. The goal of this proposal is to understand the nature and consequences of these defects, as they will provide a better understanding of the properties of cAMP signaling and to predict the outcome of pharmacological interventions targeting this signaling pathway. Genetic and biochemical approaches will be used to understand the interactions of PDE4s with 1 and 2 adrenergic receptors. The experimental plan is developed along three Specific Aims. With the first aim we will probe the role of PDE4D activation in -adrenergic signaling and their function. The second specific aim will be devoted to the analysis of the AR/PDE4 complexes and their role in receptor function. The last specific aim will probe the effect of disruption of these complexes on the function of cardiomyocytes in vitro and in vivo. In addition to shedding new light on sympathetic regulations of the cardiac function, these studies will provide rational basis for novel therapeutic strategies that target PDE and cAMP signaling. PUBLIC HEALTH RELEVANCE: The proposed experiments will shed light on the function of receptors essential for cardiac function and the role of cyclic nucleotide phosphodiesterases in the signals arising from these receptors. In view of the large number of programs in the pharmaceutical industry developing phosphodiesterase inhibitors for a variety of indications, a better understanding of the role of these targets in signaling will provide a rational basis for safer and more specific drugs. New therapeutic opportunities will open with a better understanding of the function of phosphodiesterase subtypes, as drugs affecting single isoforms may have considerable therapeutic advantages over non selective inhibitors. Finally, these studies will provide the foundation of data on which to search for human alleles of phosphodiesterase genes associated with increased risk for sudden cardiac death and other cardiovascular disorders.